Development of a simple and rapid monoclonal antibody-based flow through immunogold assay (FIA) for detection of Emphasis Type='Italic'>Aeromonas hydrophila/Emphasis>

摘要

In this study, we developed a simple, low-cost, and rapid flow through immunogold assay (FIA) for detection of Aeromonas hydrophila in fish tissues and validated. The developed assay relied on colloidal gold conjugated highly specific monoclonal antibody (B8E3) against 20?kDa protein of A. hydrophila. The assay can be completed within 5?min including antigen (sample) preparation and exhibited no cross-reaction with other major aquatic pathogens such as Vibrio anguillarum , V. parahaemolyticus , V. harveyi , white spot syndrome virus ( WSSV), and Aphanomyces invadans. A wider range of Aeromonas species were tested including A. caviae , A. dhakensis , A. sobria , A. veronii , A. culicicola , A. sharmana , and A. schubertii and no cross-reactivity could be observed. The developed technique FIA could detect 10_(3)?CFU/ml of aeromonad cells. In addition, the FIA is as sensitive as the first-step polymerase chain reaction (PCR) and 1000 times sensitive than the immunodot blot method. The validation study revealed the accuracy of FIA by comparing with the standard first-step PCR. The proposed FIA is low-cost, easy-to-use, and compatible for field-level analysis of A. hydrophila in fish tissues. Our developed FIA could successfully detect A. hydrophila at early stage of the infection and could be easily availed by farmers, fish producers, non-technicians, and technicians in the aquaculture industry.